Chromatography is a set of techniques for separating a mixture into its constituents. For instance, in a liquid chromatography application, a pumping system takes in and delivers a mixture of liquid solvents to an autosampler, where an injected sample awaits its arrival. In an isocratic chromatography application, the composition of the liquid solvents remains unchanged, whereas in a gradient chromatography application, the solvent composition varies over time. The mobile phase and the injected sample, which is dissolved in a mixture of solvents, passes to a column, referred to as the stationary phase. By passing the mixture through the column, the various components in the sample separate from each other at different rates and thus elute from the column at different times. A detector receives the separated components from the column and produces an output from which the identity and quantity of the analytes may be determined.
Preferably, the solvent composition (or composition gradient) delivered to the autosampler is a desired, stable composition. Some solvent delivery systems form the desired solvent composition by combining two or more solvents (and/or other fluids) prior to delivery of the composition to the pumping system. Such an arrangement may be referred to as a “low-pressure” gradient system because the mixing occurs on the intake (i.e., low pressure) side of the pumping system.
Before the solvent composition or gradient reaches the injector, however, and thus before injection can take place, a pre-injector dwell volume (also called the gradient delay volume) must first be delivered. In general, the pre-injector dwell volume is the volume of fluid from the point where the solvent composition or gradient forms to the injector valve. Depending upon various factors, such as the use of a mixer, pre-injector dwell volumes can be relatively large (in hundreds of microliters), and, therefore, introduce a sizeable delay to the start of each injection. Historically, however, this delay was insignificant compared to the lengthy chromatographic run times that took tens of minutes and even hours.
This delay becomes problematic, though, when run times are in terms of just a few minutes. For example, if the pre-injector dwell volume is approximately 400 μL, and the pump produces a flow rate of 350 μL per minute, then the time for the gradient to reach the point of injection is over a minute. Thus, the dead time introduced by the pre-injector dwell volume can be a significant percentage of a run, particularly when run times are only minutes in length.